title: Breakdown and release of myofilament proteins during ischemia and ischemia/reperfusion in rat hearts: identification of degradation products and effects on the pCa-force relation.
abstract: Our objective in experiments reported here was to identify myofilament proteins of rat hearts either lost or degraded by cardiac ischemia (15- or 60-minute duration) with and without 45 minutes of reperfusion. We correlated these changes with alterations in myofilament sensitivity to Ca2+ and maximum force generation. Protein degradation and loss were assessed by high-performance liquid chromatography, SDS-PAGE, Western blotting analysis, and amino acid sequencing. Compared with nonischemic control hearts, bundles of skinned fibers from hearts subjected to ischemia alone demonstrated a decrease in maximum force generation and an increase in sensitivity to Ca2+. These changes in function were increased with the duration of the ischemia and with reperfusion. With increasing duration of ischemia, there was an increased loss and degradation of myofibrillar alpha-actinin and troponin I (TnI) at its C-terminus. Alpha-actinin and TnI were most susceptible to ischemia, but with 60 minutes of ischemia/reperfusion, there was also degradation of myosin light chain-1 (MLC1) involving a clip of residues 1 to 19. The MLC1 degradation product was detected in the reperfusion effluent (along with troponin T, tropomyosin, and alpha-actinin) but not in the tissue with 60 minutes of ischemia with no reperfusion. Moreover, with ischemia the following proteins became associated with the myofibrils: GAPDH and proteins of the mitochondrial ATP synthase complex. Our results provide new evidence regarding the mechanism by which ischemia/reperfusion causes myocardial injury and support the hypothesis that an important element in the injury is altered activity and structure of the myofilaments.
authors.affiliation: Department of Physiology and Biophysics, College of Medicine, University of Illinois at Chicago, USA.
authors.forename: J E
authors.initials: JE
authors.lastname: Van Eyk
authors.forename: F
authors.initials: F
authors.lastname: Powers
authors.forename: W
authors.initials: W
authors.lastname: Law
authors.forename: C
authors.initials: C
authors.lastname: Larue
authors.forename: R S
authors.initials: RS
authors.lastname: Hodges
authors.forename: R J
authors.initials: RJ
authors.lastname: Solaro
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D006652:Histological Techniques; D066298:In Vitro Techniques; D008840:Microfilament Proteins / Q000378:metabolism*; D017202:Myocardial Ischemia / Q000378:metabolism*; D015428:Myocardial Reperfusion Injury / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism*; D009994:Osmolar Concentration; D051381:Rats
chemical_list: D008840:Microfilament Proteins; D002118:Calcium